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1.
Chinese Journal of Immunology ; (12): 15-18,24, 2018.
Artigo em Chinês | WPRIM (Pacífico Ocidental) | ID: wpr-702665

RESUMO

Objective:To study the influences of Mycoplasma penumoniae capsular polysaccharide (CPS) on the phagocytosis and membrane molecules expression of the human peripheral blood mononuclear cells derived dendritic cells by binding to dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN),so as to know the effect of CPS on the maturation of dendritic cells.Methods:M.pneumoniae strain was cultivated and CPS was extracted.Human peripheral blood mononuclear cells were separated and induced to dendritic cells,then identified the cells by flow cytometry and observation under the microscope.CPS was used to treat dendritic cells or cells pretreated with DC-SIGN monoclonal antibody,and then FITC-dextran phagocytosis and surface markers CD83,HLA-DR,CD80 and CD86 were detected by flow cytometry.Results:The dendritic cells tended to form colony groups.The positive rate of CD11c molecule in the cultured dendritic cells was about 86.27%.After stimulated by CPS,the FITC-dextran fluorescence mean intake by dendritic cells were increased (P<0.05),while the cell surface membrane molecules CD83,HLA-DR,CD80 and CD86 were decreased significantly when compared with the PBS treated control cells(P<0.05).When blocked DC-SIGN with the monoclonal antibody,the FITC-dextran fluorescence mean and membrane molecules expression had no statistical difference with the control cells(P>0.05).Conclusion:M.pneumoniae CPS can promote the phagocytic function of DC and inhibit the expression of CD83,HLA-DR,CD80 and CD86.

2.
Mol Pharm ; 14(3): 649-657, 2017 03 06.
Artigo em Inglês | MEDLINE | ID: mdl-28211700

RESUMO

How genotypic variation results in phenotypic differences is still a challenge for biology. In the field of drug metabolism, the means by which specific cytochrome P4502D6 (CYP2D6) genotypes yield different phenotypes at various levels (molecular, cellular, and organismal) is an important question, as differences in CYP2D6 activity can contribute to adverse drug reactions. Herein, the genotype of CYP2D6 was determined along with the absolute content of CYP2D6 and microsomal protein per gram of liver in human liver microsomes, the molecular, cellular (microsomal, tissue, organ), and organismal phenotype of CYP2D6 determined; the effect of genotype on each phenotype of CYP2D6-mediated dextromethorphan clearance (CL) was delineated, and the overall genotype-phenotype relationship for CYP2D6 was charted. We demonstrate that changes in the cellular and organismal CL phenotypes are markedly greater than changes seen at the molecular level. With individuals carrying the 1661CC polymorphism, for example, the most noticeable change took place in organ CL phenotype (4.17-fold), followed by tissue (3.75-fold), organism (3.69-fold), microsomal (3.09-fold), and molecular (1.66-fold) phenotypes. In addition, the biggest intragenotype individual coefficient of variation in organismal phenotype was observed in the 1661GG individuals, which reached 104.5%, followed by that of 100TT, 100CT, 1661GC, 100CC, and 1661CC polymorphisms (102.7%, 62.4%, 53.5%, 49.7%, and 44.8%, respectively). Our study has allowed us to chart the genotype-phenotype relationship for CYP2D6 from the molecular to the organismal level as well as allowed us to determine intragenotype individual variation in phenotype with each genotype.


Assuntos
Citocromo P-450 CYP2D6/genética , Dextrometorfano/metabolismo , Taxa de Depuração Metabólica/genética , Variação Genética/genética , Genótipo , Humanos , Cinética , Fenótipo
3.
Drug Metab Dispos ; 44(8): 1193-200, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27271371

RESUMO

Human cytochrome P450 oxidoreductase (POR) provides electrons for all microsomal cytochromes P450 (P450s) and plays an indispensable role in drug metabolism catalyzed by this family of enzymes. We evaluated 100 human liver samples and found that POR protein content varied 12.8-fold, from 12.59 to 160.97 pmol/mg, with a median value of 67.99 pmol/mg; POR mRNA expression varied by 26.4-fold. POR activity was less variable with a median value of 56.05 nmol/min per milligram. Cigarette smoking and alcohol consumption clearly influenced POR activity. Liver samples with a 2286822 TT genotype had significantly higher POR mRNA expression than samples with CT genotype. Homozygous carriers of POR2286822C>T, 2286823G>A, and 3823884A>C had significantly lower POR protein levels compared with the corresponding heterozygous carriers. Liver samples from individuals homozygous at 286823G>A, 1135612A>G, and 10954732G>A generally had lower POR activity levels than those from heterozygous or wild-type samples, whereas the common variant POR*28 significantly increased POR activity. There was a strong association between POR and the expression of P450 isoforms at the mRNA and protein level, whereas the relationship at the activity level, as well as the effect of POR protein content on P450 activity, was less pronounced. POR transcription was strongly correlated with both hepatocyte nuclear factor 4 alpha and pregnane X receptor mRNA levels. In conclusion, we have elucidated some potentially important correlations between POR single-nucleotide polymorphisms and POR expression in the Chinese population and have developed a database that correlates POR expression with the expression and activity of 10 P450s important in drug metabolism.


Assuntos
Sistema Enzimático do Citocromo P-450/metabolismo , Fígado/enzimologia , Consumo de Bebidas Alcoólicas/genética , Consumo de Bebidas Alcoólicas/metabolismo , China , Sistema Enzimático do Citocromo P-450/genética , Bases de Dados Factuais , Regulação Enzimológica da Expressão Gênica , Frequência do Gene , Fator 4 Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/metabolismo , Heterozigoto , Homozigoto , Humanos , Isoenzimas , Microssomos Hepáticos/enzimologia , Polimorfismo de Nucleotídeo Único , Receptor de Pregnano X , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Fumar/genética , Fumar/metabolismo
4.
Oncotarget ; 7(31): 50612-50623, 2016 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-27203676

RESUMO

The lack of information concerning individual variation in drug-metabolizing enzymes is one of the most important obstacles for designing personalized medicine approaches for hepatocellular carcinoma (HCC) patients. To assess cytochrome P450 (CYP) in the metabolism of endogenous and exogenous molecules in an HCC setting, the activity changes of 10 major CYPs in microsomes from 105 normal and 102 HCC liver tissue samples were investigated. We found that CYP activity values expressed as intrinsic clearance (CLint) differed between HCC patients and control subjects. HCC patient samples showed increased CLint for CYP2C9, CYP2D6, and CYP2E1 compared to controls. Meanwhile, CYP1A2, CYP2C8, and CYP2C19 CLint values decreased and CYP2A6, CYP2B6, and CYP3A4/5 activity was unchanged relative to controls. For patients with HCC accompanied by fibrosis or cirrhosis, the same activity changes were seen for the CYP isoforms, except for CYP2D6 which had higher values in HCC patients with cirrhosis. Moreover, CYP2D6*10 (100C>T), CYP2C9*3 (42614 A>C), and CYP3A5*3 (6986A>G) polymorphisms had definite effects on enzyme activities. In the HCC group, the CLint of CYP2D6*10 mutant homozygote was decreased by 95% compared to wild-type samples, and the frequency of this homozygote was 2.8-fold lower than the controls.In conclusion, the activities of CYP isoforms were differentially affected in HCC patients. Genetic polymorphisms of some CYP enzymes, especially CYP2D6*10, could affect enzyme activity. CYP2D6*10 allelic frequency was significantly different between HCC patients and control subjects. These findings may be useful for personalizing the clinical treatment of HCC patients as well as predicting the risk of hepatocarcinogenesis.


Assuntos
Carcinoma Hepatocelular/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Neoplasias Hepáticas/metabolismo , Adulto , Idoso , Citocromo P-450 CYP2C9/metabolismo , Citocromo P-450 CYP2D6/metabolismo , Citocromo P-450 CYP2E1/metabolismo , Citocromo P-450 CYP3A/metabolismo , Feminino , Fibrose/tratamento farmacológico , Frequência do Gene , Genótipo , Homozigoto , Humanos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Neoplasias Hepáticas/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Mutação , Polimorfismo Genético , Fumar
5.
Bioorg Med Chem Lett ; 22(6): 2350-3, 2012 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-22365754

RESUMO

Four new compounds, oliganthins A-D (1-4), and one known caged xanthone gaudichaudione H (5) were isolated from the stems of Garcinia oligantha. The structures of the new compounds were elucidated by spectroscopic evidences. All of the five compounds were evaluated for their apoptosis-inducing effects using HeLa-C3 cells which have been genetically engineered to produce a fluorescent biosensor capable of detecting caspase-3 activation. All of them induced cell apoptosis at 10 µM or lower concentrations. The apoptotic activity of oliganthins A, B and gaudichaudione H were further confirmed by detecting the cleavage of PARP, which is the substrate of activated caspase-3, in these compounds-treated cells using the method of Western blot. Moreover, the values of IC(50) were measured for all five compounds on HeLa cells using the MTT assay. Among them, gaudichaudione H had the lowest IC(50) value of 0.90 µM, while the other four new compounds had IC(50) values of 1.58, 1.52, 4.15, and 7.82 µM, respectively. These results show that gaudichaudione H has the strongest apoptosis-inducing effect and cell growth inhibition effect among these xanthones and it may have the potential to be developed into a new anticancer agent.


Assuntos
Antineoplásicos Fitogênicos/química , Apoptose/efeitos dos fármacos , Garcinia/química , Xantenos/química , Xantonas/química , Antineoplásicos Fitogênicos/isolamento & purificação , Antineoplásicos Fitogênicos/farmacologia , Western Blotting , Caspase 3/genética , Caspase 3/metabolismo , Feminino , Expressão Gênica , Genes Reporter , Células HeLa , Humanos , Espectroscopia de Ressonância Magnética , Extratos Vegetais/química , Caules de Planta/química , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/patologia , Xantenos/isolamento & purificação , Xantenos/farmacologia , Xantonas/isolamento & purificação , Xantonas/farmacologia
6.
Chinese Medical Journal ; (24): 753-757, 2004.
Artigo em Inglês | WPRIM (Pacífico Ocidental) | ID: wpr-284919

RESUMO

<p><b>BACKGROUND</b>Daxx has been identified as a nuclear protein that involves in apoptosis and transcriptional repression. Daxx co-localizes with the promyelocytic leukemia (PML) protein and regulates transcription. Human Daxx (hDaxx) is a protein that functions as a transcriptional regulation through its interaction with some DNA-associated proteins. The aim of this study was to explore the transcriptional regulatory effect of hDaxx interacting with adenovirus (Ad) 12 E1B (Ad12E1B) 55-kDa oncoprotein.</p><p><b>METHODS</b>The co-localization of hDaxx-Ad12E1B or hDaxx-PML protein in the nucleus was observed under a confocal microscope. Interaction of hDaxx and Ad12E1B was analyzed by yeast two-hybrid assay. Direct binding of hDaxx and Ad12E1B was analyzed using coimmunoprecipitation and Western blot in vivo and in vitro. The activity of a luciferase reporter gene, which was regulated by an hDaxx modulated thymidine kinase (TK) promoter, was detected in an automat luminometer.</p><p><b>RESULTS</b>Ad12E1B, which co-localized with hDaxx in the nuclei of G401-CC3 cells, disrupted the co-localization of hDaxx and PML in the PML oncogenic domains (PODs). hDaxx bound directly to Ad12E1B in vivo and in vitro. hDaxx interacted with Ad12E1B along its full length. Ad12E1B enhanced transcriptional repression activity of hDaxx.</p><p><b>CONCLUSION</b>Ad12E1B disrupts the co-localization of hDaxx with PML in PODs and enhances transcriptional repression activity of hDaxx.</p>


Assuntos
Humanos , Proteínas Adaptadoras de Transdução de Sinal , Proteínas E1B de Adenovirus , Fisiologia , Proteínas de Transporte , Genética , Linhagem Celular Tumoral , Peptídeos e Proteínas de Sinalização Intracelular , Proteínas de Neoplasias , Proteínas Nucleares , Genética , Proteína da Leucemia Promielocítica , Proteínas Repressoras , Fisiologia , Fatores de Transcrição , Transcrição Gênica , Proteínas Supressoras de Tumor
7.
Chinese Medical Journal ; (24): 997-1001, 2004.
Artigo em Inglês | WPRIM (Pacífico Ocidental) | ID: wpr-284862

RESUMO

<p><b>BACKGROUND</b>This study was designed to investigate the potential pathogenicity of Mycoplasma penetrans (M. penetrans) and its molecular mechanisms responsible for the induction of iNOS gene expression in mouse macrophages stimulated by lipid-associated membrane proteins (LAMPs) prepared from M. penetrans.</p><p><b>METHODS</b>Mouse macrophages were stimulated with M. penetrans LAMPs to assay the production of nitric oxide (NO). The expression of inducible nitric oxide synthase (iNOS) was detected by RT-PCR and Western blotting. The activity of nuclear factor kappaB (NF-kappaB) and the effects of pyrrolidine dithiocarbamate (PDTC), an inhibitor of NF-kappaB, on the production of nitric oxide and the expression of iNOS were also assessed in mouse macrophages treated with M. penetrans LAMPs by indirect immunofluorescence and Western blotting.</p><p><b>RESULTS</b>M. penetrans LAMPs stimulated mouse macrophages to produce nitric oxide in a dose- and time-dependent manner. The mRNA and protein levels of iNOS were also upregulated in response to LAMP stimulation and inhibited by PDTC treatment. M. penetrans LAMPs were found to trigger NF-kappaB activation, a possible mechanism for the induction of iNOS expression.</p><p><b>CONCLUSION</b>This study demonstrated that M. penetrans may be an important etiological factor of certain diseases due to the ability of M. penetrans LAMPs to stimulate the expression of iNOS, which is probably mediated through the activation of NF-kappaB.</p>


Assuntos
Animais , Camundongos , Proteínas de Bactérias , Farmacologia , Células Cultivadas , Indução Enzimática , Lipoproteínas , Farmacologia , Proteínas de Membrana , Farmacologia , Mycoplasma penetrans , Química , NF-kappa B , Metabolismo , Óxido Nítrico , Óxido Nítrico Sintase , Óxido Nítrico Sintase Tipo II , RNA Mensageiro
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